The platelet cytoskeleton contains elements of the prothrombinase complex

Triton-insoluble cytoskeletons prepared from thrombin-activated platelets were found to potentiate the activation of prothrombin (prothrombinase activity). Cytoskeletons prepared from red cells or lymphoblasts contained no prothrombinase activity. The platelet prothrombinase activity was dependent on cytoskeletal-associated Factor Va, and exogenously added Factor Xa and prothrombin. Cytoskeletons contained 38% of the total platelet prothrombinase activity. Both platelets and cytoskeletons displayed half-maximal activities at similar prothrombin concentrations. The role of lipids in the cytoskeletal prothrombinase activity was investigated. Cytoskeletons were found to contain 3.8% of the total platelet phospholipids, consisting of the following lipids expressed as percentage of total present in platelets: 6.0% sphingomyelin, 3.8% phosphatidylcholine, 2.9% phosphatidyl-ethanolamine, 4.4% phosphatidylinositol, and 2.2% phosphatidylserine. The cytoskeletal prothrombinase activity and the lipid phosphorus content of cytoskeletons decreased after treatment of cytoskeletons with various doses of phospholipase C. Incubation of cytoskeletons with the highest concentrations tested (10 micrograms/ml) resulted in a 72% loss of phosphatidylserine and 84% loss of cytoskeletal prothrombinase activity. Cytoskeletal prothrombinase activity destroyed by phospholipase C treatment could be restored to control levels by treatment of hydrolyzed cytoskeletons with total cytoskeletal lipid or mixtures of phosphatidylserine/phosphatidylcholine (25:75% by weight). These results suggest that the cytoskeletal prothrombinase complex in addition to containing Factor Va, as has been previously shown (15), contains a lipid cofactor activity consisting in part of phosphatidylserine.     

The M/SAR elements of the bithorax complex in Drosophila melanogaster

The bithorax (BX) complex of Drosophila is a complex polygenic region with a multifactorial system of regulation. One of the levels of the regulatory system of the BX complex is its association with the nuclear skeleton structures through a specific interaction of the M/SAR DNA with the nuclear matrix proteins. In the present work, M/SAR elements were mapped on the molecular-genetic map of the region. All of the elements examined were found to colocalize with regulatory elements and form clusters that restrict/bracket the genetically active domains. All M/SAR DNA revealed was shown to bins specifically to the purified Drosophila melanogaster lamin.     

Gene order in the major histocompatibility complex of the rat

The loci in the major histocompatibility complex (MHC) of the rat which code for class I and class II antigens—RT1.A and RT1.B, respectively — have previously been separated by laboratory-derived recombinants and by observations in inbred and wild rats. Closely linked to the MHC is the growth and reproduction complex (Grc) which contains genes influencing body size (dw-3) and fertility (ft). These phenotypic markers were used in this study to orient the A and B loci of the MHC. Two recombinants were used for mapping. The BIL(R1) animal is a recombinant between the MHC and Grc, and it carries the haplotype RT1.A ^lB^lGrc⁺. The r10 animal is an intra-MHC recombinant, and it has the haplotype RT1.A ⁿB¹ Grc. These recombinants were characterized serologically, by mixed lymphocyte reactivity, by immune responsiveness to poly (Glu⁵²Lys³³Tyr¹⁵) and by the presence of the dw-3 gene. The data demonstrate that the gene order of the loci is: dw-3-RT1.B-RT1.A.     

Age-related changes of elements in renal arteries of Thai and Japanese and the relationships among elements

To examine whether there were differences between reces in regard to age-related changes of elements and the relationships among elements in the arteries, the authors investigated both the renal arteries of Thai and Japanese. The Thai subjects consisted of 27 men and 11 women, ranging in age from 27 to 88 yr, whereas the Japanese subjects consisted of 19 men and 26 women, ranging in age from 61 to 99 yr. After the ordinary dissections at Chiang Mai University and Nara Medical University were completed, the bilateral renal arteries were resected and the element contents were determined by inductively coupled plasma-atomic emission spectrometry. In the Thai, a slight accumulation of calcium and phosphorus occurred in the fifties, but thereafter headly increased. In contrast, in the Japanese, an accumulation of calcium and phosphorus began to occur in the seventies and increased markedly in the eighties. The result reveraled that a higher accumulation of calcium and phosphorus occurred in the renal arteries of the Japanese in old age compared with those of the Thai. Regarding the relationships among elements, extremely or very significant direct correlations were found among the contents of calcium, phosphorus, magnesium, and sodium in both the renal arteries of the Thai and Japanese, except for magnesium and sodium contents in the renal arteries of the Thai.     

Predicting rules on organization of cis-regulatory elements, taking the order of elements into account

MOTIVATION: In eukaryotes, rules regarding organization of cis-regulatory elements are complex. They sometimes govern multiple kinds of elements and positional restrictions on elements. RESULTS: We propose a method for detecting rules, by which the order of elements is restricted. The order restriction is expressed as element patterns. We extract all the element patterns that occur in promoter regions of at least the specified number of genes. Then, we find significant patterns based on the expression similarity of genes with promoter regions containing each of the extracted patterns. When we applied our method to Saccharomyces cerevisiae, we detected significant patterns overlooked by previous methods, thus demonstrating the utility of our method for analyses of eukaryotic gene regulation. We also suggest that several types of element organization exist: (i) those in which only the order of elements is important, (ii) order and distance both are important and (iii) only the combination of elements is important. AVAILABILITY: The program for extracting element patterns is available upon request.     

Alu elements of the primate major histocompatibility complex

The chromosomal region constituting the major histocompatibility complex (MHC) has undergone complex evolution that is often difficult to decipher. An important aid in the elucidation of the MHC evolution is the presence of Alu elements (repeats) which serve as markers for tracing chromosomal rearrangements. As the first step toward the establishment of sets of evolutionary markers for the MHC, Alu elements present in selected MHC haplotypes of the human species, the gorilla, and the chimpanzee were identified. Restriction fragments of cosmid clones from the libraries of the three species were hybridized with Alu-specific probes, Alu elements were amplified by the polymerase chain reaction, and the amplification products were sequenced. In some cases, sequences of the regions flanking the Alu elements were also obtained. Altogether, 31 new Alu elements were identified, representing six Alu subfamilies. The average density of Alu elements in the MHC is one element per four kilobases (kb) of sequence. Alu elements have apparently been inserted steadily into the MHC over the last 65 million years (my). On average, one Alu element is inserted into the primate MHC every 4 my. Analysis of the human DR3 haplotype supports its origin by duplication from an ancestral haplotype consisting of DRB1 and DRB2 genes. The sharing of an old Alu element by the DRB1 and DRB2 genes, in turn, supports their divergence from a common ancestor more than 55 my ago.     

Temporal order and the evolution of complex acoustic signals

The evolution of complex signals may be favoured by hidden preferences or pre-existing sensory biases. Females of two species of grey treefrogs (Hyla chrysoscelis and Hyla versicolor) were tested with combinations of a conspecific advertisement call and acoustic appendages. Appendages consisted of aggressive calls and segments of advertisement calls from conspecific males and males of three other species and bursts of filtered noise. When a wide variety of these acoustic appendages followed the advertisement call, the resulting compound signal was often more attractive than the same advertisement call alone. When the same appendages led advertisement calls, however, the compound signal was never more attractive and sometimes less attractive. The order effect was especially strong in tests of H. versicolor in which advertisement-call duration was decreased. These results cannot be explained by a general pre-existing bias for extra stimulation per se. Rather, order and other effects may constrain the evolution and subsequent modification of complex and extravagant signals, examples of which have been reported for a wide range of taxa.     

Oligosaccharide order in a membrane-incorporated complex glycosphingolipid.

Galactosylceramide (GalCer) and the ganglioside, GM1, were 2H-labelled at C-6 (the hydroxymethyl moiety) of their single terminal galactosyl residues. Each deuterated glycosphingolipid was incorporated at a biologically relevant low concentration into multibilayers of 1-palmitoyl-2-oleoylphosphatidylcholine (POPC). 2H-NMR spectra of aqueous dispersions of GalCer-POPC in the liquid crystal phase were characteristic of restricted headgroup motion (ordering) with effective axial symmetry. The degree of headgroup ordering was analogous to that of GalCer in pure aqueous multibilayers (Skarjune, R. and Oldfield, E. (1979) Biochim. Biophys. Acta 556, 208-218). In the case of GM1, 2H-labelled in the terminal galactose residue of the pentasaccharide headgroup, the 2H-NMR spectra were remarkably like those of the simple glycolipid, GalCer. This suggests substantial restriction of motion about the glycosidic and sugar-ceramide bonds of the complex GM1 headgroup, and that both lipids have comparable degrees of orientational averaging (fluctuation) about the bilayer normal. The result is the first direct demonstration that headgroup orientational order can exist for a complex glycolipid incorporated into 'fluid' bilayer membranes. Such behaviour argues for the possibility of modulation of membrane receptor properties through surface effects on average headgroup orientation and conformation.     

The complex of xylan and iodine: the induction and detection of nanoscale order

The complex of xylan and iodine and its formation in a solution of xylan, CaCl₂, and I₂ + KI was investigated by UV/Vis, second-derivative UV/Vis, and Raman spectroscopy. The complex forms only at very high concentrations of CaCl₂, suggesting that when the water available in the solution is not sufficient to fully hydrate the calcium cation the chelation with the hydroxyl groups of the xylan can occur. The electronic spectra indicate that iodine is present in the form of three linear polyiodides I₉³⁻, I₁₁³⁻, and I₁₃³⁻ structures, which the Raman spectra show to be linear aggregates of the I₃⁻ and I₅⁻ substructures. Iodide concentration has a significant influence on the relative population of I₉³⁻, I₁₁³⁻, and I₁₃³⁻, as well as I₃⁻ and I₅⁻, which lead to changes in both the UV/Vis absorption maxima shifts and changes in the Raman spectra. The key difference between this system of complexes with the linear polyiodide aggregates and that of amylose is that the longest aggregate observed with the amylose system, the I₁₅³⁻ polyanion, is not observed with the xylans. This indicates that the ordered arrays in the xylan-iodine complex do not exceed 4 nm in length. It is not possible to conclude at this time whether the ordered segment of the xylan molecule is linear or helical. If it is linear the length of the longest ordered arrays would be eight xylose residues. The number would exceed eight if the xylan molecule were helically wound.     

Bringing order to a complex molecular machine: The assembly of the bacterial flagella

The bacterial flagellum is an example of elegance in molecular engineering. Flagella dependent motility is a widespread and evolutionarily ancient trait. Diverse bacterial species have evolved unique structural adaptations enabling them to migrate in their environmental niche. Variability exists in the number, location and configuration of flagella, and reflects unique adaptations of the microorganism. The most detailed analysis of flagellar morphogenesis and structure has focused on Escherichia coli and Salmonella enterica. The appendage assembles sequentially from the inner to the outer-most structures. Additionally the temporal order of gene expression correlates with the assembly order of encoded proteins into the final structure. The bacterial flagellar apparatus includes an essential basal body complex that comprises the export machinery required for assembly of the hook and flagellar filament. A review outlining the current understanding of the protein interactions that make up this remarkable structure will be presented, and the associated temporal genetic regulation will be briefly discussed.     

Characterization of new regulatory elements within the Drosophila bithorax complex

The homeotic Abdominal-B (Abd-B) gene expression depends on a modular cis-regulatory region divided into discrete functional domains (iab) that control the expression of the gene in a particular segment of the fly. These domains contain regulatory elements implicated in both initiation and maintenance of homeotic gene expression and elements that separate the different domains. In this paper we have performed an extensive analysis of the iab-6 regulatory region, which regulates Abd-B expression at abdominal segment A6 (PS11), and we have characterized two new polycomb response elements (PREs) within this domain. We report that PREs at Abd-B cis-regulatory domains present a particular chromatin structure which is nuclease accessible all along Drosophila development and both in active and repressed states. We also show that one of these regions contains a dCTCF and CP190 dependent activity in transgenic enhancer-blocking assays, suggesting that it corresponds to the Fab-6 boundary element of the Drosophila bithorax complex.     

Insertion events of CR1 retrotransposable elements elucidate the phylogenetic branching order in galliform birds

Using standard phylogenetic methods, it can be hard to resolve the order in which speciation events took place when new lineages evolved in the distant past and within a short time frame. As an example, phylogenies of galliform birds (including well-known species such as chicken, turkey, and quail) usually show low bootstrap support values at short internal branches, reflecting the rapid diversification of these birds in the Eocene. However, given the key role of chicken and related poultry species in agricultural, evolutionary, general biological and disease studies, it is important to know their internal relationships. Recently, insertion patterns of transposable elements such as long and short interspersed nuclear element markers have proved powerful in revealing branching orders of difficult phylogenies. Here we decipher the order of speciation events in a group of 27 galliform species based on insertion events of chicken repeat 1 (CR1) transposable elements. Forty-four CR1 marker loci were identified from the draft sequence of the chicken genome, and from turkey BAC clone sequence, and the presence or absence of markers across species was investigated via electrophoretic size separation of amplification products and subsequent confirmation by DNA sequencing. Thirty markers proved possible to type with electrophoresis of which 20 were phylogenetically informative. The distribution of these repeat elements supported a single homoplasy-free cladogram, which confirmed that megapodes, cracids, New World quail, and guinea fowl form outgroups to Phasianidae and that quails, pheasants, and partridges are each polyphyletic groups. Importantly, we show that chicken is an outgroup to turkey and quail, an observation which does not have significant support from previous DNA sequence- and DNA-DNA hybridization-based trees and has important implications for evolutionary studies based on sequence or karyotype data from galliforms. We discuss the potential and limitations of using a genome-based retrotransposon approach in resolving problematic phylogenies among birds.     

New elements in the structure of binding sites of glucocorticoid-receptor complex in hormone-regulated genes

An analysis of the structure of DNA sites responsible for binding to glucocorticoid-receptor complex (GlRC) was carried out. The use of the frequency matrices and of a variant of the perception method made it possible to establish that in the GlRC binding site on both sides of the known conservative nucleotide sequence (nucleus) there were additional conservative elements which seemed to be able to modulate the efficiency of GlRC binding. A criterion is worked out for detecting the potential GlRC binding sites in given sequences. It is based on the simultaneous use of several perceptron matrices. The efficiency of detection of GlRC binding sites by means of the proposed criterion is by an order higher than that performed according to the GlRC binding site consensus (Beato et al. [2]).     

Concentration ratios of stable elements for selected biota in Japanese estuarine areas

For the estimation of radiation doses to organisms, concentration ratios (C _R s) of radionuclides are required. In the present study, C _R s of various elements were obtained as analogues of radionuclides for algae, molluscs, and crustaceans, in eight estuarine areas around Japan. The elements measured were Na, Mg, K, Ca, V, Mn, Fe, Co, Ni, Cu, Rb, Sr, Y, Mo, Cd, La, Ce, Pr, Nd, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu, Pb, and U. The geometric mean (GM) values of C _R s (GM-C _R s) for alkali and alkaline earth elements, Mo, and U for all biota, as well as V for crustaceans, were less than 100 L/kg, while GM-C _R s for the other elements were higher. When the obtained GM-C _R s were compared with the C _R s recommended in IAEA Technical Report Series 422 for marine organisms, no big differences between them were found; however, several elements (i.e. Cd and U for algae, Mn for molluscs, and Pb for crustaceans) were lower than the recommended C _R s. In the present study, conversion factors (the ratio of C _R for the whole body to that for muscle) for molluscs and crustaceans were also calculated, since data on edible parts of these organisms are generally available in the literature. For crustaceans, GM conversion factors of all the elements were more than one. For molluscs, GM conversion factors of rare earth elements and U were slightly higher than those for crustaceans, while GM conversion factors of the other elements were almost the same and less than 10. These results indicate that some elements tend to be concentrated in the internal organs of biota collected in the estuarine areas. For environmental radiological assessment, conversion factors from tissue to whole-body C _R values are useful parameters.